DESCRIPTION (Applicant's Abstract): This proposal, "In vivo Spatio-Temporal Optical Imaging of Focal Ischemia", is submitted in response to the NINDS program announcement for Mentored Quantitative Research Career Development Awards (K25). The candidate seeks to gain training in the neurobiology of focal cerebral ischemia in order to apply advanced optical imaging techniques to the study of cerebral ischemia in animal models. Since tissue in the ischemic penumbra can potentially be salvaged, it is important to quantify the blood flow changes and molecular events within the penumbra in vivo. In this proposal high resolution optical imaging techniques will be used to assess pathophysiologic events in the penumbra in vivo following focal ischemia. Specifically, the hemodynamic and cellular events that lead to alterations in the microvasculature or apoptotic cell death will be examined following focal ischemic brain injury. The spatial and temporal characteristics of the blood flow changes in the transition from core to penumbra to normal tissue in a rat model of focal ischemia will be quantified in vivo using two-dimensional laser speckle blood flow imaging. The spatial and temporal alterations in blood flow in the ischemic penumbra in response to eNOS upregulation will be dynamically measured. Having characterized the blood flow changes in the penumbra, the level of caspase-3 activity, which is an indicator of apoptosis, will be measured in vivo in mice overexpressing recombinant GFP-based caspase substrates with two-photon fluorescence microscopy. This unique animal model will allow in vivo visualization and quantification of the effects of a caspase inhibitor (DEVD-fmk) on the level of caspase-3 cleavage throughout the ischemic penumbra. Through the training that this award provides, the candidate will develop the knowledge of neuroscience and stroke physiology necessary to lead an independent research career applying advanced optical imaging techniques to study the ischemic penumbra.